Laing N G, Siddique T, Bartlett R, Yamaoka L H, Hung W Y, Pericak-Vance M A, Roses A D
Department of Medicine, Duke University Medical Center, Durham, NC.
Clin Genet. 1989 Jun;35(6):393-8. doi: 10.1111/j.1399-0004.1989.tb02963.x.
DNA isolated from a family segregating a deletion in the Duchenne muscular dystrophy gene and control families was digested with restriction enzymes, Southern transferred, and probed with a radioactive dystrophin cDNA probe. The resulting autoradiographs were analyzed with a densitometric spectrophotometer to detect carriers of the deletion. The carrier status of females in the deletion pedigree was independently determined by genomic probes and confirmed by densitometry. In many Duchenne families, deletions will only be observed using cDNA probes which show few restriction fragment length polymorphisms (RFLPs). Such deletions would normally have to be detected using dosage gels. The spectrophotometric densitometry technique used by us does not require dosage gels, and avoids problems arising from non-informative meioses and cross-overs. It should be possible to screen every family with an exon deletion by spectrophotometric densitometry provided the presently available cDNA is suitably reduced to produce fewer bands on autoradiographs.
从一个杜氏肌营养不良基因存在缺失的家系以及对照家系中分离出的DNA，用限制性内切酶消化，进行Southern印迹转移，并用放射性肌营养不良蛋白cDNA探针进行杂交。用密度分光光度计分析所得的放射自显影片，以检测缺失的携带者。缺失家系中女性的携带者状态通过基因组探针独立确定，并通过密度测定法进行确认。在许多杜氏家系中，只有使用显示很少限制性片段长度多态性（RFLP）的cDNA探针才能观察到缺失。这种缺失通常必须使用剂量凝胶来检测。我们使用的分光光度密度测定技术不需要剂量凝胶，并且避免了由无信息减数分裂和交叉产生的问题。只要将目前可用的cDNA适当减少，以在放射自显影片上产生更少的条带，就应该能够通过分光光度密度测定法对每个存在外显子缺失的家系进行筛查。
